This program will complete research and development, and launch implementation and commercialization of a technology for differential diagnosis of infectious diseases that integrates PCR and atmospheric pressure chemical ionization mass spectroscopy (MS) into a stable and sensitive digital assay platform with the advantages of simultaneous detection and discrimination of multiple targets. This platform, Mass Tag PCR will first be used to distinguish pathogens implicated in acute, severe respiratory disease including SARS CoV and influenza viruses, and later extended to other NIAID priority agents. Features that will enhance the probability of success include: expertise of applicants in clinical microbiology, bioinformatics, and molecular diagnostics; a novel technology for mass spectrometric detection of PCR amplification products; banks of viral primer sequences, cDNA targets and mass tags; access to miniaturized mass spectrometric instruments suited to field applications; partnership in an international surveillance network wherein clinical samples can be shared for assay optimization and validation, and a commercial network for quality controlled manufacture and global distribution of reagents for molecular biology. Aim 1: Establish multiplex PCR/MS assay on conventional MS instruments for detection of respiratory pathogens that cause severe pneumonia; Aim 2: Build portable APCI MS instruments to support multiplex PCR/MS platform; Aim 3: Transfer multiplex assays from conventional to portable APCI MS instruments; Aim 4: Establish PCR/MS technology at network laboratories; Aim 5: Transfer technology to detection of NIAID Category A, B, and C priority agents.